Melon aphid-borne yellows virus (MABYV), a member of the genus Polerovirus in the family Solemoviridae, has widely spread in recent years and cause yellowing disease on cucurbits. Here, we obtained the complete genome sequence of MABYV bottle guard (Lagenaria siceraria) isolate MABYV-KF, and constructed its infectious cDNA clone under the control of the cauliflower mosaic virus (CaMV) 35S promoter by Gibson assembly. The 5,677 nt of its genome shared more than 94.00% sequence identity with the two known MABYV isolates. The inoculation results showed that MABYV infectious cDNA clone could systemically infect bottle guard, cucumber and muskmelon plants, and cause typical yellowing symptom. The virus progeny from the infectious clone could be transmitted between bottle guard plants by aphid. Further analyses revealed that point mutations in the F-box-like motif (Pro57) and C-terminal conserved sequence (Phe211) of P0 cause low viral accumulations in systematic leaves and failed to induce symptom. The infectious clone will be potentially a tool in the investigation of viral pathogenesis, virus-virus interaction and virus-host/-vector interactions.

Swine could serve as a natural reservoir for a large variety of viruses, including potential zoonotic enteric viruses. The presence of viruses with high genetic similarity between porcine and human strains may result in the emergence of zoonotic or xenozoonotic infections. Furthermore, the globalization and intensification of swine industries exacerbate the transmission and evolution of zoonotic viruses among swine herds and individuals working in swine-related occupations. To effectively prevent the public health risks posed by zoonotic swine enteric viruses, designing, and implementing a comprehensive measure for early diagnosis, prevention, and mitigation, requires interdisciplinary a collaborative ''One Health" approach from veterinarians, environmental and public health professionals, and the swine industry. In this paper, we reviewed the current knowledge of selected potential zoonotic swine enteric viruses and explored swine intensive production and its associated public health risks.

Autophagy is a lysosomal degradative pathway responsible for recycling cytosolic proteins and organelles and also functions as an innate defense mechanism that host cells use against viral infection. While many viruses have evolved mechanisms to antagonize the antiviral effects of the autophagy pathway, others subvert autophagy to facilitate replication. For flaviviruses, both the positive and negative role of autophagy in virus replication has been reported. The interplay between autophagy and tick-borne encephalitis virus (TBEV) in innate immune cells is largely unknown. Here we report the relationship between an autophagy and TBEV replication in mouse macrophage cell line PMJ2-R using Hypr strain of TBEV. First, we examined the effect of Hypr infection on the autophagy pathway. We detected a mild and a temporary increase of autophagy marker LC3-II in Hypr-infected cells. The role of autophagy in TBEV replication was evaluated in autophagy related gene 5 (Atg5) knockdown cells (shAtg5). Our results showed that during an early stage of Hypr infection the viral titers were increased, while later on, at 72 hpi, the titers have declined in shAtg5 cells compared to control. Moreover, the higher number of virus-positive cells was observed in shAtg5 cells in early stage of infection and correlated with enhanced virus entry. Finally, we found an increased production of IFN-β in Hypr-infected shAtg5 cells in comparison to control at 48 and 72 hpi implicating that autophagy restricts the amount of IFN produced by TBEV-infected macrophages. To conclude, in mouse macrophages TBEV replication is controlled by autophagy in time dependent manner, having temporally an antiviral and then a pro-viral role during infection. Our study points out to a delicate and complex involvement of autophagy machinery at level of virus entry and IFN-β production when controlling TBEV infection.

Channel catfish virus (CCV), an important member of the family Alloherpesviridae, causes a lethal infection in channel catfish. As with most animal viruses, the initial step of infection by CCV is entry into host cells, which is also a promising antiviral target for CCV disease. This study investigated the mechanism of host cell invasion by CCV using a series of biochemical inhibitor assays in channel catfish cells. CCV infection in host cells was does-dependently inhibited when cells were treated with endosomal acidification inhibitors (5 μM chloroquine, 50 nM bafilomycin A1, and 1 mM ammonium chloride) and hypertonic medium (50 mM sucrose) , which suggests that CCV invades host cells in a manner dependent on low-pH and the endocytic pathway. Moreover, when the cells were pretreated with inhibitors of clathrin-mediated endocytosis, including chlorpromazine (2 μM) and dynasore (50 μM), the CCV infection in the host cells was strongly inhibited. In contrast, the destruction of cellular cholesterol by methyl-β-cyclodextrin and nystatin and inhibition of macropinocytosis had no effect on viral entry. Altogether, these findings indicate that CCV infects host cells via clathrin-mediated endocytosis in a low-pH-dependent manner, suggesting that this CCV entry pathway offers an antiviral target against CCV disease.

At the end of 2019, in China, clinical signs and symptoms of unknown etiology have been reported in several patients whose sample sequencing revealed pneumonia caused by the SARS-CoV-2 virus. COVID-19 is a disease triggered by this virus, and in 2020, the World Health Organization declared it a pandemic. Since then, efforts have been made to find effective therapeutic agents against this disease. Identifying novel natural antiviral drugs can be an alternative to treatment. For this reason, antimicrobial peptides secreted by anurans' skin have gained attention for showing a promissory antiviral effect. Hence, this review aimed to elucidate how and which peptides secreted by anurans' skin can be considered therapeutic agents to treat or prevent human viral infectious diseases. Through a literature review, we attempted to identify potential antiviral frogs' peptides to combat COVID-19. As a result, the Magainin-1 and -2 peptides, from the Magainin family, the Dermaseptin-S9, from the Dermaseptin family, and Caerin 1.6 and 1.10, from the Caerin family, are molecules that already showed antiviral effects against SARS-CoV-2 in silico. In addition to these peptides, this review suggests that future studies should use other families that already have antiviral action against other viruses, such as Brevinins, Maculatins, Esculentins, Temporins, and Urumins. To apply these peptides as therapeutic agents, experimental studies with peptides already tested in silico and new studies with other families not tested yet should be considered.

The rabies infection is a zoonotic viral disease in humans and is spread by both wild and domestic carnivores. This study aimed to molecularly characterize the field strains of the rabies virus circulating in Turkey between 2013 and 2020. Brain samples obtained from 16 infected animals (eight cattle, one donkey, three foxes, three dogs, and one marten) were tested. Full nucleoprotein (N) and glycoprotein (G) gene sequences were used to determine the genetic and antigenic characteristics of the rabies virus field strains. The phylogenetic analyses revealed that the 16 field strains identified in Turkey belonged to the Cosmopolitan lineage.

Autographa californica multiple nucleocapsid polyhedrosis virus (AcMNPV) genome contains several homologous regions (hrs) which can be used as origins of viral DNA replication and transcriptional enhancers. Early studies have found that some hrs can not only stimulate viral early promoters, but also enhance gene expression driven by viral late promoters. In this study, by adding hr4a to the upstream of different types of viral promoters and enhanced green fluorescent protein (egfp) reporter gene, we investigated the enhancement effect of hr4a on eGFP expression. The results showed that hr4a increased the gene transcription controlled by early promoters, but had no enhancer effect on late promoters.

COVID-19 caused by SARS-CoV-2 virus has had profound impact on the world in the past two years. Intense research is going on to find effective drugs to combat the disease. Over the past year several vaccines were approved for immunization. But SARS-CoV-2 being an RNA virus is continuously mutating to generate new variants, some of which develop features of immune escape. This raised serious doubts over the long-term efficacy of the vaccines. We have identified a unique mannose binding plant lectin from Narcissus tazetta bulb, NTL-125, which effectively inhibits SARS-CoV-2 replication in Vero-E6 cell line. In silico docking studies revealed that NTL-125 has strong affinity to viral Spike RBD protein, preventing it from attaching to hACE2 receptor, the gateway to cellular entry. Binding analyses revealed that all the mutant variants of Spike protein also have stronger affinity for NTL-125 than hACE2. The unique α-helical tail of NTL-125 plays most important role in binding to RBD of Spike. NTL-125 also interacts effectively with some glycan moieties of S-protein in addition to amino acid residues adding to the binding strength. Thus, NTL-125 is a highly potential antiviral compound of natural origin against SARS-CoV-2 and may serve as an important therapeutic for management of COVID-19.

Erratum in Virus Res. 2022 Sep;318:198828.

Hepatitis B virus (HBV) infection is a major cause of hepatocellular carcinoma (HCC) worldwide. HBV X protein (HBx) is potentially the most oncogenic among HBV-encoding proteins, while HBV integration, which is frequently observed in HCC, contributes to HCC development. However, the molecular mechanism underlying HBV-induced hepatocarcinogenesis remains unclear. In this study, we identified the fusion HBx, the HBx-human fusion protein derived from HBV integrant, in Hep3B cells and investigated its role in hepatocarcinogenesis. The identified full-length fusion mRNA was 3,725 bp in length, and the fusion HBx, which consisted of 1-140 amino acids of HBx followed by 61 amino acids from the human genome, was translated from the fusion mRNA. The fusion HBx knockdown resulted in reduced cell proliferation and invasion, and loss of tumor development in nude mice. Moreover, the fusion HBx, but not wild HBx, provided anchorage-independent growth ability in soft agar although its transactivation ability was abrogated. Microarray analysis revealed that fusion HBx deregulated endoplasmic reticulum (ER) stress response by modifying ATF3, ATF4, and ATF6 transcription. Interestingly, the effects of fusion HBx on ER stress signaling pathway were similar to those of C-terminal truncated HBx, but significantly different from those of wild HBx. Our findings suggest that the fusion HBx plays a significant role in hepatocarcinogenesis by modifying ER stress response and could be an attractive target for the treatment of HBV-induced HCC.

Escherichia coli, a bacterium that causes severe foodborne diseases, is transmitted to humans primarily through the consumption of contaminated foods. These foodborne pathogens are causing a public health problem that requires alternative control approaches, such as bacteriophage (phage) biocontrol. In this study, we characterized vB_EcoM_Tw01 (vTw01) isolated from sewage and vB_EcoM_Tcm05 (vTcm05) isolated from chicken meat. Both vTw01 and vTcm05 were assigned to the family Myoviridae based on their morphology, with the former exhibiting a narrow host range with low minimum inhibitory multiplicity of infection (miMOI), and the latter exhibiting a broad host range with high miMOI. The latent periods of these phages were 20 and 30 min for vTw01 and vTcm05, while the burst sizes were ∼140 and ∼300 PFU/cell, respectively. They were relatively stable over a wide range of pH values and temperatures. The bioinformatics analysis of the genomic sequence suggests that vTw01 and vTcm05 have double-stranded DNA with genome sizes of 170,107 bp and 149,059 bp, respectively. Bacteriophage encoded enzymes, such as tail-lysozyme, spanin Rz, holin, cell wall hydrolase (CWH), and endolysin, were identified in the genome of both phages. In conclusion, this study investigated the morphological, physiological, and genomic features of two E. coli phages isolated from different sources. It was confirmed that these phages and their enzymes can serve as potential candidates for phage biocontrol.

BACKGROUND: Emergence of new variant of SARS-CoV-2, namely omicron, has posed a global concern because of its high rate of transmissibility and mutations in its genome. Researchers worldwide are trying to understand the evolution and emergence of such variants to understand the mutational cascade events. METHODS: We have considered all omicron genomes (n = 302 genomes) available till 2nd December 2021 in the public repository of GISAID along with representatives of variants of concern (VOC), i.e., alpha, beta, gamma, delta, and omicron; variant of interest (VOI) mu and lambda; and variant under monitoring (VUM). Whole genome-based phylogeny and mutational analysis were performed to understand the evolution of SARS CoV-2 leading to emergence of omicron variant. RESULTS: Whole genome-based phylogeny depicted two phylogroups (PG-I and PG-II) forming variant specific clades except for gamma and VUM GH. Mutational analysis detected 18,261 mutations in the omicron variant, majority of which were non-synonymous mutations in spike (A67, T547K, D614G, H655Y, N679K, P681H, D796Y, N856K, Q954H), followed by RNA dependent RNA polymerase (rdrp) (A1892T, I189V, P314L, K38R, T492I, V57V), ORF6 (M19M) and nucleocapsid protein (RG203KR). CONCLUSION: Delta and omicron have evolutionary diverged into distinct phylogroups and do not share a common ancestry. While, omicron shares common ancestry with VOI lambda and its evolution is mainly derived by the non-synonymous mutations.

Brazil ranks as third in terms of total number of reported SARS-CoV-2 cases globally. The COVID-19 epidemic in Brazil was characterised by the co-circulation of multiple variants as a consequence of multiple independent introduction events occurring through time. Here, we describe the SARS-CoV-2 variants that are currently circulating and co-circulating in the country, with the aim to highlight which variants have driven the different epidemic waves. For this purpose, we retrieved metadata information of Coronavirus sequences collected in Brazil and available at the GISAID database. SARS-CoV-2 lineages have been identified along with eleven variants, labelled as VOCs (Alpha, Gamma, Beta, Delta and Omicron) VOIs (Lambda and Mu) VUMs (B.1.1.318) and FMVs (Zeta, Eta and B.1.1.519). Here we show that, in the Brazilian context, after 24 months of sustained transmission and evolution of SARS-CoV-2, local variants (among them the B.1.1.28 and B.1.1.33) were displaced by recently introduced VOCs firstly with the Gamma, followed by Delta and more recently Omicron. The rapid spread of some of those VOCs (such as Gamma and Omicron) was also mirror by a large increase in the number of cases and deaths in the country. This in turn reinforces that, due to the emergence of variants that appear to induce a substantial evasion against neutralizing antibody response, it is important to strengthen genomic effort within the country and how vaccination still remains a critical process to protect the vulnerable population, still at risk of infection and death.

Viral metagenomic analysis of wisteria leaf sample in Iran detected one dicistrovirus: aphid lethal paralysis virus (ALPV). The complete genome sequence of ALPV-Ir-Wi was 9824 nucleotides (nt) in length (excluding the 3'-poly(A) tail), and contained two ORFs, an intergenic untranslated region of 197 nt flanked by a 538 nt 5' UTR and a 576 nt 3' UTR. Comparison with 21 other ALPV genomic sequences from different parts of the world revealed that it most closely resembled the Turkish and Israeli isolates. Pairwise identity analysis showed significant variability in genome sequences among ALPV isolates with genomic nucleotide identities of 78.35-99.15%. In addition to codon mutations, insertions/deletions and recombination also contributed to genetic variability. To explore the genetic variation and molecular evolution of ALPV, ORF2 gene sequences of 18 non-recombinant isolates were analyzed. The isolates belonged to two principal clades (FST=0.614) and showed a considerable genetic diversity (0.140±0.01). Most populations were polyphyletic, indicating that they had not been isolated long enough to reach reciprocal monophyly. There was no significant correlation between genetic and geographical distances or host origins. Pairwise FST and Nm values showed a meaningful differentiation and relatively infrequent gene flow between two compared populations (the Middle East vs. East Asia, the Middle East vs. Africa), and moderate gene flow for East Asian and African populations. Genes in the ALPV genome were subject to strong purifying selection during evolution, and most codons were under negative selection or neutral evolution. The results indicated a relatively stable and conserved genomic composition with a low codon usage bias in all of the assayed ALPV coding sequences. Recombination, natural selection, gene flow, and founder effects were found to be the main evolutionary factors that can affect the genetic structure of ALPV populations.

BACKGROUND: Hepatitis B virus-related hepatocellular carcinoma (HBV-HCC) will contribute to more than half of the liver deaths worldwide. This study aimed to investigate the prognostic value of TSPEAR-AS1 in HBV-HCC and its role in the HBV-HCC progression. METHODS: HBV-HCC tissue and adjacent non-cancerous tissues (ANT) were detected to figure out the expression level of TSPEAR-AS1 using real-time quantitative PCR. The relationship between TSPEAR-AS1 expression and each important clinical characteristic was evaluated. And the prognostic significance of TSPEAR-AS1 was assessed by Kaplan-Meier curve and Cox regression analysis. CCK-8 and Transwell assays were performed to observe the effects of TSPEAR-AS1 on HBV-HCC cell proliferation, migration, and invasion. RESULTS: The TSPEAR-AS1 expression was downregulated in HBV-HCC tissues, as well as in HBV-HCC cell lines. The downregulation of TSPEAR-AS1 showed a significant association with TNM stage, clinical stage, and vascular invasion and predicted poor prognosis of HBV-HCC patients. Overexpression of TSPEAR-AS1 inhibited HBV-HCC cell ability of proliferation, migration, and invasiveness. TSPEAR-AS1 may bind to miR-1915-5p in HCC. CONCLUSION: TSPEAR-AS1 expression was downregulated in HBV-HCC and may serve as a potential prognostic factor. TSPEAR-AS1 might exert a suppressor role in HBV-HCC through inhibiting tumor cell proliferation, migration, and invasion.

E. coli O157: H7 is known as a high-risk food-born pathogen, and its removal is vital for maintaining food safety. The increasing trend of food-borne diseases caused by this bacterium and other pathogens indicates the low efficiency of the methods to remove pathogens from foodstuffs. One of the new and effective methods is to use of a bio-control agent called bacteriophage, which has shown good function in eliminating and reducing pathogens. In this study, a novel bacteriophage was isolated and identified from the slaughterhouse wastewater to control E. coli O157: H7. This bacteriophage belonged to the Myoviridae family. Two bacterial genera including E. coli and Salmonella, were allocated to determine the bacteriophage host range; the result showed that the anti- Salmonella effect of phage was low. The phage was stable at high temperature (80 °C) and caused an acceptable reduction in the E. coli O157: H7 (4.18 log CFU / mL for 10 h). The isolated bacteriophage was corroborated to be completely safe based on the whole genome sequencing and lack of any virulence factor from the host bacteria. Considering the characteristics of this phage and its function in vitro, this bacteriophage may be used as an effective bio-control agent in foods with the possible E. coli O157: H7 -induced contamination.

Vaccination prevents and reduces the severity of influenza virus infections. Continuous evolution of influenza hemagglutinin (HA) and neuraminidase (NA) supports the virus to evade pre-existing immunity, which demands vaccines to be reformulated every year. Incorporation of polymerase basic protein 1 (PB1) viral RNA (vRNA) of the same origin of HA and NA vRNA has been observed in previous pandemic viruses and occasionally reported for influenza A vaccine prototype strains of prior seasons. At this point, it remains to be explored whether this phenomenon translates into an improved growth phenotype. In this work, we showed that the HA vRNA of A(H1N1)pdm09 is generally incorporated with the PB1 vRNA of the same origin, establishing the beneficial effect of the presence of PB1 and the pattern of the PB1-HA co-incorporation in the A(H1N1)pdm09 model. We further investigated the putative interplay between PB1 and antigenic proteins regarding the vRNA composition of the progeny and observed that vRNA segregation does not appear to be mainly determined by protein-protein interactions; while vRNA-vRNA interactions can be suggested as the main driving force. Our data also indicate an increase in the hemagglutination capacity and neuraminidase activity due to incorporation of PB1, HA and NA from A(H1N1)pdm09, in comparison with the recombinant virus incorporating only HA and NA from A(H1N1)pdm09 - which have the potential to improve current limitations regarding antigenicity and immunogenicity of influenza vaccines. Further knowledge of the complex vRNA-vRNA interaction network between PB1 and HA will additionally contribute to improve current vaccine formulation, and to gradually optimize the production of A(H1N1)pdm09 reverse genetics vaccine seed virus towards a higher cost-effectiveness.

Avian influenza viruses (AIVs) circulating in wild ducks are rarely transmitted directly to chickens. Previous studies demonstrated that chickens possess fucosylated and/or sulfated α2,3 sialosides on their tracheal epithelia, whereas intestinal epithelia of ducks express canonical α2,3 sialosides. Turkeys, the third major poultry species in the world, are known to show broad susceptibility to various avian influenza viruses. To elucidate the molecular basis of the broad susceptibility of turkeys to duck and chicken AIVs, we characterized various receptors for AIVs on their tissues. The experimental infection of turkeys demonstrated their dual susceptibility to duck and chicken AIVs. Further, comprehensive histochemical analyses using lectins, anti-glycan antibodies, and recombinant hemagglutinins, combined with glycosidase digestions, identified the presence of fucosylated and/or sulfated in addition to canonical α2,3 sialosides on their respiratory epithelia. The receptor distributions in turkeys were consistent with their dual susceptibility to duck and chicken AIVs. Also, our findings suggested the potential roles of turkeys in interspecies transmission of AIVs from ducks to chickens.

High genetic diversity of porcine reproductive and respiratory syndrome virus (PRRSV) isolates is a major obstacle for the development of effective PRRS vaccines. A chimeric highly pathogenic PRRSV2 (HP-PRRSV2) strain containing the consensus sequence of ORF2-6 genes was constructed in our previous study, which could induce broadly neutralizing antibodies (bnAbs) and confer satisfied cross protection against virulent NADC30-like isolate. To further elucidate the roles of minor and major envelope proteins encoded by ORF2-4 and ORF5-6 genes in conferring cross protection, two chimeric HP-PRRSV2 strains (rJS-ORF2-4-CON and rJS-ORF5-6-CON) containing consensus sequences of ORF2-4 or ORF5-6 were constructed and rescued in this study. The rJS-ORF5-6-CON strain has similar replication efficiency as the backbone HP-PRRSV2 rJSTZ1712-12 virus, while rJS-ORF2-4-CON has significantly lower in vitro and in vivo replication efficiency comparing to rJS-ORF5-6-CON. Animal inoculation indicated that both rJS-ORF2-4-CON and rJS-ORF5-6-CON did not cause obvious clinical signs in piglets and could induce heterologous nAbs after immunization. Challenge with a virulent heterologous NADC30-like SD17-38 isolate showed that even though both immunized groups presented lower viremia, faster virus elimination, less fever and alleviated lung gross lesions when compared with the only challenged pigs, rJS-ORF2-4-CON and rJS-ORF5-6-CON could not confer enough cross protection. Considering the bnAbs and satisfied cross protection induced by the chimeric virus containing ORF2-6 consensus sequence, our results support that minor and major envelope proteins play synergistic roles in inducing broader nAbs and conferring better cross protection.

Studies are needed to better understand the genomic evolution of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). This study aimed to describe viral quasispecies population of upper and lower respiratory tract by next-generation sequencing in patients admitted to intensive care unit. A deep sequencing of the S gene of SARS-CoV-2 from 109 clinical specimens, sampled from the upper respiratory tract (URT) and lower respiratory tract (LRT) of 77 patients was performed. A higher incidence of non-synonymous mutations and indels was observed in the LRT among minority variants. This might be explained by the ability of the virus to invade cells without interacting with ACE2 (e.g. exploiting macrophage phagocytosis). Minority variants are highly concentrated around the gene portion encoding for the Spike cleavage site, with a higher incidence in the URT; four mutations are highly recurring among samples and were found associated with the URT. Interestingly, 55.8% of minority variants detected in this locus were T>G and G>T transversions. Results from this study evidenced the presence of selective pressure and suggest that an evolutionary process is still ongoing in one of the crucial sites of spike protein associated with the spillover to humans.

NADC30-like porcine reproductive and respiratory syndrome virus (PRRSV), which is highly homologous to the NADC30 strain isolated in the United States. The NADC30-like PRRSV was first reported in 2014 in China, where it spread and gradually caused an epidemic. Currently, growing research has shown that NADC30-like strains have greater propensity to recombine with other PRRSV strains, particularly the PPRSV vaccine virus used clinically, making the prevention and control of PRRSV highly complex. To carry out an in-depth molecular biology and virulence analysis, a full-length infectious clone of the NADC30-like strain was successfully constructed and rescued by reverse genetic manipulation. The rescued virus, rZJqz, was indistinguishable from its parental virus, ZJqz21, based on virological characteristics. Further animal experiments demonstrated that rZJqz retained similar pathogenicity and induced the typical clinical symptoms and viral shedding observed in the ZJqz21 challenge model. Together, these results provide a useful tool for further study of the biological characteristics and pathogenicity of NADC30-like strains. Moreover, these findings also provide a solid foundation for studying the recombination of different PRRSVs and developing new and effective universal vaccines in the future.

Spring viraemia of carp virus (SVCV) poses a serious threat to aquaculture industry due to the lack of approved antiviral treatments. Therefore, a novel arctigenin derivative, 4-(2-methylimidazole) octanoxy-arctigenin (MON), was synthesized to assess the antiviral activity against SVCV in vitro and in vivo. The results indicated MON decreased the SVCV glycoprotein (G) gene expression in vitro by a maximum inhibitory rate of > 99% at 3.5 μM. Furthermore, MON showed the protective effect on epithelioma papulosum cyprinid (EPC) cells and considerably decreased the cytopathic effect (CPE). More importantly, MON inhibited SVCV G gene expression levels in vitro at the half-maximal activity (IC50) of 0.18 μM at 48 h. For in vivo studies, MON demonstrated anti-SVCV activity by enhancing the survival rate of zebrafish (Danio rerio) after infection via pelvic fin base injection. These results tended to be consistent with MON decreasing the SVCV titer of infected zebrafish. During this time, viral loads of the spleen and kidney have declined in SVSV-infected zebrafish. Based on the histopathological assay, MON exhibited the high protective effect in the spleen and kidney of SVCV-infected fish. Combined, MON is on track to become a novel agent to address SVCV infection in aquaculture.

Halyomorpha halys (Stål, 1855) (Hemiptera: Pentatomidae), the brown marmorated stink bug, is an invasive pentatomid native to East-Asia, and introduced worldwide in recent times. It is a polyphagous pest with approximately 300 host plants, which, due to its plasticity, reproductive and feeding behavior, long-distance flight, and walking as well as human-mediated dispersal ability, is able to cause significant economic and ecological damage. In several cases pest control mediated by insecticide treatments leads to unsatisfactory efficacy, mostly due to insect recovery ability. Thus, the most promising method for the long-term management of this pest has been focused with growing emphasis on classical biological control strategies. In this framework, viruses have untill now been poorly investigated in H. halys with only a single virus described from the US territory. For this reason we investigated the virome associated with a small and well described population of H. halys from Piedmont (Italy) describing for the first time 7 new viral sequences belonging to different taxonomical groups. Further studies will be necessary to assess the biological and ecological effects the viruses have on their host. Due to the agricultural importance of this insect, the biological characterization of these viruses would give important information on the possibility to exploit viral entities as biological control agents. Finally, the presence of a such relevant number of viruses from a small population suggests a wide association between the brown marmorated stink bug and viral entities. Further studies to determine the possible exploitation of viral sequences to trace different populations are ongoing.

Enterobacter cloacae is an opportunistic pathogen that widely distributes in various environments, often causing hospital and community-acquired infections. The limited phage resources and poor genomic data hinder the understanding of the lysis mechanism of Enterobacter cloacae phages. In the present study, we characterized a newly discovered lytic phage PZJ0206 that infects an E. cloacae strain isolated from sewage water into the Pearl River in Guangdong, China. The morphological characteristics of the icosahedral head and short tail of PZJ0206 were observed by transmission electron microscopy. The one-step growth curve showed that PZJ0206 has a relatively short latent period and an average burst size of approximately 124 PFU (plaque forming unit)/infected cell at an MOI of 0.1. The genome of phage PZJ0206 consists of 40,526 bp with a GC content of 48.62% and contains 45 putative open reading frames (ORFs). Based on genomic characteristics, comparative genomics and phylogenetic analysis, it was strongly demonstrated that PZJ0206 is a novel member of the genus Berlinvirus in the family Autographiviridae. Our work provides basic information on Enterobacter cloacae phage PZJ0206 and lays the foundation for the further study of the evolution of phage PZJ0206 and interactions between the phage and its host.

Porcine reproductive and respiratory syndrome; Porcine reproductive and respiratory syndrome virus; PRRS; Whole genome sequencing; Quasispecies; WUR allele.

The black armyworm Spodoptera cosmioides is a pest of increasing importance in Cry1Ac-Bt toxin crops and non-Bt crops of soybean and cotton in Brazil. Here we characterized a baculovirus isolated from extracts of S. cosmioides that died with symptoms of nuclear polyhedrosis. The putative novel virus exhibited polyhedral occlusion bodies (OBs) with virions containing multiple rod-shaped nucleocapsids, characteristic of alphabaculoviruses. The virus isolate was named Spodoptera cosmioides nucleopolyhedrovirus isolate CNPSo-72 (SpcoNPV-CNPSo-72). SpcoNPV-CNPSo-72 was lethal to third-instar S. cosmioides caterpillars but not to S. frugiperda under the tested viral concentrations. Moreover, SpcoNPV-CNPSo-72 contained a circular 147,763 bp long genome and a G + C content of 44.8% with 151 annotated ORFs (10 unique for baculovirus) and five homologous regions (hrs). The 38 currently defined baculovirus core genes were found in the SpcoNPV-CNPSo-72 genome. After phylogenetic analysis, the novel virus was found to be closely related to other members of Alphabaculovirus, especially to the Spodoptera-infecting viruses, which included Spodoptera eridania nucleopolyhedrovirus isolate 251, Spodoptera litura nucleopolyhedrovirus isolate II, Spodoptera exigua multiple nucleopolyhedrovirus isolate US-1, Spodoptera eridania nucleopolyhedrovirus isolate CNPSo-165, and Spodoptera frugiperda multiple nucleopolyhedrovirus isolate 19. Surprisingly, the new baculoviral genome was found to code for a putative arginine-associated tRNA gene with a predicted intronic sequence of 105 nt. The gene was found inside the bjdp CDS. Overall, baculoviruses are pathogens that lethally infect insect larvae and their study allows a better understanding of large DNA virus evolution, which provides important insights for the development and improvement of biological control agents.

The emergence of the Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2) represents an unprecedented threat for the human population, necessitating rapid and effective intervention measures. Given the main infection route by airborne transmission, significant attention has been bestowed upon the use of antiseptic mouthrinses as a way to possibly reduce infectious viral titers. However, clinical evaluations are still sparse. Thus, we evaluated a wide variety of antiseptic agents that can be used as mouthrinses for their antiviral effects in vitro and their respective mode of action. One of the most promising antiseptic agents (benzalkoniumchloride, BAC) was used in a randomized placebo-controlled clinical trial with subsequent analysis of viral loads by RT-qPCR and virus rescue in cell culture. Mechanistic analysis revealed that treatment with BAC and other antiseptic agents efficiently inactivated SARS-CoV-2 in vitro by primarily disrupting the viral envelope, without affecting viral RNA integrity. However, the clinical application only resulted in a mild reduction of viral loads in the oral cavity. These results indicate that gargling with mouthrinses comprising single antiseptic agents may play a minor role towards a potential reduction of transmission rates and thus, these findings are of utmost importance when considering alternative COVID-19 prevention strategies.

To provide insights into the biology of the attenuated canine distemper virus (CDV) Onderstepoort (OP) strain (large plaque forming variant), design next-generation multivalent vaccines, or further investigate its promising potential as an oncolytic vector, we employed contemporary modifications to establish an efficient OP-CDV-based reverse genetics platform. Successful viral rescue was obtained however only upon recovery of a completely conserved charged residue (V13E) residing at the N-terminal region of the large protein (L). Although L-V13 and L-V13E did not display drastic differences in cellular localization and physical interaction with P, efficient polymerase complex (P+ L) activity was recorded only with L-V13E. Interestingly, grafting mNeonGreen to the viral N protein via a P2A ribosomal skipping sequence (OPneon) and its derivative V-protein-knockout variant (OPneon-Vko) exhibited delayed replication kinetics in cultured cells. Collectively, we established an efficient OP-CDV-based reverse genetics system that enables the design of various strategies potentially contributing to veterinary medicine and research.